Programming Switchable Transcription of Topologically Constrained DNA

TLDR: It is found that TO-DNA maintains the transcription activity whereas the activity is critically dependent on the configuration of the T7 promoter in a folded DNA nanostructure, and the design of TO-DNAs provides a means for shape-dependent gene delivery, enriching the toolbox of genetic engineering and synthetic biology.

Abstract: Genomic DNA is compacted via chromatin condensation in mammalian cells, and transcription of such topologically constrained DNA to messenger RNA is under strict spatiotemporal regulation. Nevertheless, control of DNA topology has been poorly explored in in vitro transcription and gene transfection. Here we report the construction of topologically ordered (TO-) prokaryotic genes composed of linear DNA templates appended with a T7 promoter sequence with the use of DNA self-assembly. We find that TO-DNA maintains the transcription activity whereas the activity is critically dependent on the configuration of the T7 promoter in a folded DNA nanostructure. By prescribing the position and the intactness of the T7 promoter, we can dynamically activate or repress transcription in response to specific DNA key strands in a Boolean logic manner. Bioorthogonal switchable transcription is realized with the insertion of multiple genes in a TO-DNA. Further, implementing TO-DNA in living bacteria leads to switchable transcription of fluorescent RNA aptamers for light-up cell imaging. Hence, the design of TO-DNAs provides a means for shape-dependent gene delivery, enriching the toolbox of genetic engineering and synthetic biology.